Reactivity of natural and induced human antibodies to MUC1 mucin with MUC1 peptides andn-acetylgalactosamine (GalNAc) peptides

Abstract

Antibodies (Abs) to MUC1 occur naturally in both healthy subjects and cancer patients and can be induced by MUC1 peptide vaccination. We compared the specificity of natural and induced MUC1 Abs with the objective of defining an effective MUC1 vaccine for active immunotherapy of adenocarcinoma patients. Serum samples, selected out of a screened population of 492 subjects for their high levels of IgG and/or IgM MUC1 Abs, were obtained from 55 control subjects and from 26 breast cancer patients before primary treatment, as well as from 19 breast cancer patients immunized with MUC1 peptides coupled to keyhole limpet hemocyanin (KLH) and mixed with QS‐21. The samples were tested with enzyme‐linked immunoassays for reactivity with (1) overlapping hepta‐ and 20‐mer peptides spanning the MUC1 tandem repeat sequence; (2) two modified 60‐mer peptides with substitutions in the PDTR (PDTA) or in the STAPPA (STAAAA) sequence of each tandem repeat; and (3) four 60‐mer glycopeptides with each 1, 2, 3 and 5 mol N‐acetylgalactosamine (GalNAc) per repeat. More than one minimal epitopic sequence could be defined, indicating that Abs directed to more than one region of the MUC1 peptide core can coexist in one and the same subject. The most frequent minimal epitopic sequence of natural MUC1 IgG and IgM Abs was RPAPGS, followed by PPAHGVT and PDTRP. MUC1 peptide vaccination induced high titers of IgM and IgG Abs predominantly directed, respectively, to the PDTRPAP and the STAPPAHGV sequences of the tandem repeat. Natural MUC1 Abs from breast cancer patients reacted more strongly with the N‐acetylgalactosamine (GalNAc) peptides than with the naked 60‐mer peptide, while reactivity with the GalNAc‐peptides was significantly reduced (2‐tailed p < 0.0001) in the MUC1 IgG and IgM Abs induced by MUC1 peptide vaccination. Whereas in cancer patients glycans appear to participate in epitope conformation, the epitope(s) recognized by MUC1 Abs induced by peptide vaccination are already masked by minimal glycosylation. Therefore, our results indicate that a MUC1 glycopeptide would be a better vaccine than a naked peptide. Int. J. Cancer 86:702–712, 2000. © 2000 Wiley‐Liss, Inc.

Cancer Biomarkers Defined by Autoantibody Signatures to Aberrant O-Glycopeptide Epitopes

“…The MUC1 mucin is heavily O-glycosylated in a large 20 amino acid tandem repeated region, which is aberrantly glycosylated in cancer (20) and can be detected in serum of late stage cancer patients (21;22). Man is in general immunologically tolerant to the MUC1 tandem repeat protein core and its normal glycosylated forms (20;23), although, several studies have shown that up to 10–15% of healthy controls and cancer patients have antibody reactivity to MUC1 peptides as detected by ELISA assays; however, the detected levels are very low and have led to contradictory results by different investigators (24–27). We recently identified an immunodominant O-glycopeptide epitope (Tn/STn-MUC1) that results from aberrant glycosylation and for which there does not appear to be immunological tolerance (28–30) (See Supplementary Fig.…”

Immunology of O-Glycosylated Proteins: Approaches to the Design of a MUC1 Glycopeptide-Based Tumor Vaccine

“…These effects are of particular relevance in case of MUC1, since in contradiction to previous assumptions this mucin seems to be more densely O-glycosylated on tumor cells compared to normal cells, though with truncated glycans related to the Tn and T antigens and their sialylated derivatives [11]. It is not surprising, accordingly, that natural and experimentally induced antibody responses in humans are more directed to glycosylated MUC1 epitopes [12]. On the other hand, antibodies generated to the non-glycosylated MUC1 repeat peptide were largely unable to bind the natural, tumor-associated glycoform [12].…”

“…It is not surprising, accordingly, that natural and experimentally induced antibody responses in humans are more directed to glycosylated MUC1 epitopes [12]. On the other hand, antibodies generated to the non-glycosylated MUC1 repeat peptide were largely unable to bind the natural, tumor-associated glycoform [12]. So, cancer immunologists end up with the challenge to design a MUC1-based cancer vaccine that, on the one hand, induces immune responses to the natural, heavily O-glycosylated target, but on the other, allows for active processing and MHC presentation of glycopeptides to finally elicit strong and long lasting responses to the glycotopes on tumor cells.…”

Proteomic Analysis, Immune Dysregulation, and Pathway Interconnections with Obesity

“…These self-molecules trigger B cell activation usually because of aberrant expression, mislocalization, sequencing errors 107,108 , and/or altered post-translational modifications 109 . Thus, autoantibodies can lend interesting insights into disease processes.…”